Project Summary This proposal seeks competitive renewal of my GM049369, which has been supporting our basic research in regulated gene expression by RNA binding proteins (RBPs) in the past two decades. We will focus on two families of RBPs, SR proteins and RBFoxs, which have been best characterized for their functions in regulated pre-mRNA splicing. However, our recent studies show that they are also directly involved in transcriptional control through interacting with regulatory RNAs in mammalian cells. Built on these paradigm- shifting discoveries, we propose to test a series of hypotheses by pursuing four specific aims. The first is to elucidate synergistic interactions of SR proteins in the cell to test the hypothesis that SR proteins are engaged in network interactions to establish their specificity in interacting with cis-acting RNA elements in mammalian genomes. The second aim to determine the role of SR proteins in integrating gene expression with particular emphasis on their contribution to genome organization in the nucleus. The third aim is to dissect the mechanism for RBFox2 to regulate polycomb complex targeting in the genome through their synergistic interactions on nascent RNA. The last aim will develop the ES cell system to determine RBFox2 isoform- specific functions in guiding PRC2 to chromatin during cell lineage commitment. The grand hypothesis we will test is that RBPs are broadly involved in transcriptional control through interacting with diverse regulatory RNAs in mammalian genomes. As all genes we are studying have been tightly linked to various human diseases, particularly cancer and neurodegenerative diseases, the proposed research will not only elucidate new regulatory paradigms and mechanisms, but also provide the theoretical foundation for developing new therapeutics against challenging human diseases.